Azoxymethane  (Synonyms: AOM)

The assay for Azoxymethane-induced in vitro DNA adduct formation is performed. Briefly, microsomes (0.5 to 2.0 mg/mL) are incubated with calf thymus DNA (1 mg/mL) and Azoxymethane (200 μM) in a total volume of 1.0 mL. The assay buffer consists of 0.1 M Tris-HCl (pH 7.4), 1 mM EDTA, 20 mM MgCl₂, 0.3 M KCl, and 1.5 mM NADPH. Incubations are carried out at 37°C for 60 min in a shaking water bath. An additional 30 nM of NADPH is added after the first 30 min. The reaction is stopped by the addition of 0.5 mL of ice-cold 7.5 M ammonium acetate. DNA is then extracted for tissue homogenates. Control incubations are performed without NADPH^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

Male, 8 to 10 week old, WT-A/J, IECN-A/J, and LCN-A/J mice (8 per group) are treated with either saline or Azoxymethane (7.5 mg/kg BW, s.c.), once weekly for 3 weeks. Mice are sacrificed 6 weeks post-treatment for aberrant crypt foci (ACF) detection. The entire colon is excised. A longitudinal incision is made along the entire length of the colon, which is further cut into two equal-length segments, representing proximal and distal portions of the colon. The segments are dipped in PBS to remove fecal pellets and then kept flat between filter papers in 10% buffered formalin for at least 24 h. Subsequently, the colons are immersed in freshly prepared 0.1% methylene blue for 10 min and rinsed briefly in deionized H₂O to remove excess dye. The colon is mounted carefully on a microscope slide with the mucosal surface side up and viewed under a light microscope. The ACF in the entire mucosal surface of the colon are counted blindly and independently by two investigators and recorded^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Megaraj V, et al. Role of hepatic and intestinal p450 enzymes in the metabolic activation of the colon carcinogen azoxymethane in mice. Chem Res Toxicol. 2014 Apr 21;27(4):656-62.  [Content Brief]

  [2]. Whetstone RD, et al. Colon carcinogenesis in wild type and immune compromised mice after treatment with azoxymethane, and azoxymethane with dextran sodium sulfate. Mol Carcinog. 2016 Jul;55(7):1187-95.  [Content Brief]

  [3]. Fichera A, et al. Epidermal growth factor receptor signaling is required for microadenoma formation in the mouse azoxymethane model of colonic carcinogenesis. Cancer Res. 2007 Jan 15;67(2):827-35.  [Content Brief]